INTERACTION AND REGULATION OF THE CAENORHABDITIS-ELEGANS DEATH PROTEASE CED-3 BY CED-4 AND CED-9

In the nematode Caenorhabditis elegans, three genes, ced-3, ced-4, and ced-9, play critical roles in the induction and execution of the deat h pathway, Genetic studies have suggested that ced-9 controls programm ed cell death by regulating ced-4 and ced-3, However, the mechanism by which CED-9 controls the activities of CED-4 and the cysteine proteas e CED-3, the effector arm of the cell-death pathway, remains poorly un derstood, Immunoprecipitation analysis demonstrates that CED-9 forms a multimeric protein complex with CED-4, and CED-3 in vivo. Expression of wild-type CED-4 promotes the ability of CED-3 to induce apoptosis i n mammalian cells, which is inhibited by CED-9, The pro-apoptotic acti vity of CED-4 requires the expression of a functional CED-3 protease, Significantly, loss-of-function CED-4 mutants are impaired in their ab ility to promote CED-3-mediated apoptosis, Expression of CED-4 enhance s the proteolytic activation of CED-3, We also show that CED-9 inhibit s the formation of p13 and p15, two cleavage products of CED-3 associa ted with its proteolytic activation in vivo. Moreover, CED-9 inhibits the enzymatic activity of CED-3 promoted by CED-4. Thus, these results provide evidence that CED-4 and CED-9 regulate the activity of CED-3 through physical interactions, which may provide a molecular basis for the control of programmed cell death in C. elegans.